Atf1 Is a Target of the Mitogen-activated Protein Kinase Pmk1 and Regulates Cell Integrity in Fission Yeast

Abstract

In fission yeast, knockout of the calcineurin gene resulted in hypersensitivity to Cl(-), and the overexpression of pmp1(+) encoding a dual-specificity phosphatase for Pmk1 mitogen-activated protein kinase (MAPK) or the knockout of the components of the Pmk1 pathway complemented the Cl(-) hypersensitivity of calcineurin deletion. Here, we showed that the overexpression of ptc1(+) and ptc3(+), both encoding type 2C protein phosphatase (PP2C), previously known to inactivate the Wis1-Spc1-Atf1 stress-activated MAPK signaling pathway, suppressed the Cl(-) hypersensitivity of calcineurin deletion. We also demonstrated that the mRNA levels of these two PP2Cs and pyp2(+), another negative regulator of Spc1, are dependent on Pmk1. Notably, the deletion of Atf1, but not that of Spc1, displayed hypersensitivity to the cell wall-damaging agents and also suppressed the Cl(-) hypersensitivity of calcineurin deletion, both of which are characteristic phenotypes shared by the mutation of the components of the Pmk1 MAPK pathway. Moreover, micafungin treatment induced Pmk1 hyperactivation that resulted in Atf1 hyperphosphorylation. Together, our results suggest that PP2C is involved in a negative feedback loop of the Pmk1 signaling, and results also demonstrate that Atf1 is a key component of the cell integrity signaling downstream of Pmk1 MAPK.

The Rho1p Exchange Factor Rgf1p Signals Upstream from the Pmk1 Mitogen-activated Protein Kinase Pathway in Fission Yeast

“…The finding that the up-regulation of Pck2p and Pmk1p suppressed, at least partially, the Csp defect of rgf1⌬ mutants suggests that Pck2p signals to the cell wall integrity pathway via the MAPK cascade. It has been shown that Atf1p is phosphorylated by Pmk1p under cell wall stress, but the consequences of such activation are unknown (Takada et al, 2007). Besides its role in the cell integrity pathway, Rho1p acts by regulating the biosynthesis of ␤(1,3)-glucan and the cell wall in general, and it is also required for actin polymerization.…”

“…The MAPK module is composed of MAPKKK Mkh1p (Sengar et al, 1997), MAPKK Pek1p/Shk1p (Sugiura et al, 1999;Loewith et al, 2000), and MAPK Pmk1p/Spm1p (Toda et al, 1996b;Zaitsevskaya-Carter and Cooper, 1997). Although this module was identified several years ago, little is known about its upstream components or the phosphorylation substrates activated by the cell integrity signaling pathway (Ma et al, 2006;Takada et al, 2007). The MAPKKKs at the head of the module are often activated through phosphorylation and/or as a result of their interaction with a small GTP-binding protein of the Ras/Rho family in response to extracellular stimuli (Roux and Blenis, 2004).…”

Oxidative stress response pathways: Fission yeast as archetype

“…For instance, the Pmk1 and Sty1 MAPKs not only share regulatory links (i.e. the Pyp1, Pyp2 and Ptc1 protein phosphatases) (Takada et al, 2007), but also target the same TF (i.e. Atf1) to regulate gene expression through the cell integrity and stress response pathway respectively (Figure 3).…”

Role of the RNA-binding Protein Nrd1 and Pmk1 Mitogen-activated Protein Kinase in the Regulation of Myosin mRNA Stability in Fission Yeast

“…Leucine prototrophs were selected, and the identification of strain containing one copy of cdc4M1M2 expressed from the genomic cdc4 ϩ promoter was verified by PCR and immunoblot analysis. In vitro kinase reactions were performed as previously described (Takada et al, 2007) with some modifications. Bacterially expressed and purified GST-Nrd1 in place of Atf1-FLAG was used as a substrate.…”